SLC25A24 Antibody
Description
Introduction to SLC25A24 Antibody
SLC25A24 antibodies are immunological reagents specifically designed to recognize and bind to the SLC25A24 protein, which functions as an electroneutral antiporter in the inner mitochondrial membrane. These antibodies serve as essential tools for researchers investigating mitochondrial functions, cellular metabolism, and related pathological conditions. Available as both rabbit and mouse polyclonal antibodies, they enable the detection and characterization of SLC25A24 in various experimental contexts .
The development of these antibodies has significantly advanced our understanding of mitochondrial carrier proteins and their roles in cellular homeostasis. SLC25A24 antibodies are produced through immunization of host animals with specific SLC25A24 protein fragments or recombinant proteins, followed by purification techniques to isolate the antibodies with high specificity for the target protein .
Host Species and Antibody Types
Commercial SLC25A24 antibodies are predominantly available as polyclonal antibodies raised in either rabbit or mouse hosts. Both rabbit polyclonal and mouse polyclonal versions exhibit distinct characteristics that make them suitable for different experimental conditions . The polyclonal nature of these antibodies allows for recognition of multiple epitopes on the SLC25A24 protein, enhancing sensitivity in various applications. The host species affects compatibility with secondary detection systems and potential background issues when working with tissues from related species .
Immunogen Information
Most SLC25A24 antibodies are generated using recombinant protein fragments or full-length human SLC25A24 as immunogens. For instance:
-
Proteintech's rabbit polyclonal antibody (14669-1-AP) is generated using a specific SLC25A24 fusion protein (Ag6334)
-
Abcam's rabbit polyclonal antibody (ab221120) is produced using a recombinant fragment protein within human SLC25A24 aa 1-100
-
The mouse polyclonal antibody from antibodies-online (ABIN526192) is raised against a full-length human SLC25A24 protein (AA 1-477)
Target Protein: SLC25A24
Understanding the target protein is essential for properly utilizing and interpreting results obtained with SLC25A24 antibodies. The SLC25A24 protein serves important functions in mitochondrial transport and cellular protection.
Structure and Basic Information
SLC25A24 (Solute Carrier Family 25 Member 24) is a mitochondrial carrier protein with the following characteristics:
The observed molecular weight (48 kDa) differs slightly from the calculated weight (53 kDa), which is not uncommon for membrane proteins due to their hydrophobic nature and post-translational modifications .
Functional Role and Biological Significance
SLC25A24 plays critical roles in mitochondrial function and cellular protection:
-
It functions as an electroneutral antiporter that mediates the transport of adenyl nucleotides through the inner mitochondrial membrane .
-
Initially identified as an ATP-magnesium/inorganic phosphate antiporter, it also acts as a broad specificity adenyl nucleotide antiporter, with low activity also observed with guanyl and pyrimidine nucleotides in vitro .
-
By regulating the mitochondrial matrix adenyl nucleotide pool, it helps cells adapt to changing energetic demands and indirectly regulates adenyl nucleotide-dependent metabolic pathways .
-
SLC25A24 may play a protective role against oxidative stress-induced cell death by buffering calcium levels in the mitochondrial matrix through the formation of calcium-phosphate precipitates .
These functions highlight the importance of SLC25A24 in cellular metabolism and stress response mechanisms, making its study relevant for understanding mitochondrial physiology and pathology.
Applications and Methodologies
SLC25A24 antibodies are versatile tools used in multiple laboratory techniques that enable the detection, localization, and quantification of the SLC25A24 protein in various biological samples.
Western Blotting (WB)
Western blotting is one of the primary applications for SLC25A24 antibodies, allowing for protein detection and semi-quantitative analysis:
| Antibody | Recommended Dilution | Validated Samples | Source |
|---|---|---|---|
| Proteintech 14669-1-AP | 1:1000-1:6000 | HEK-293 cells, mouse heart tissue, human placenta tissue | |
| Abcam ab221120 | 0.04-0.4 μg/ml | RT4 and U251 MG cell lysates |
Western blotting protocols typically involve heat-mediated antigen retrieval and detect the SLC25A24 protein at approximately 48 kDa . The successful detection in multiple species (human, mouse, rat) demonstrates the cross-reactivity of these antibodies, making them valuable for comparative studies .
Immunohistochemistry (IHC)
Immunohistochemistry applications allow for the visualization of SLC25A24 protein within tissue sections:
The ability to localize SLC25A24 within tissue sections provides valuable information about its distribution in different cell types and potential functional roles in specific tissues .
Immunofluorescence (IF/ICC)
Immunofluorescence techniques offer high-resolution imaging of SLC25A24 localization at the subcellular level:
Immunofluorescence studies with SLC25A24 antibodies typically reveal the expected mitochondrial localization pattern, confirming the specificity of the antibodies and providing insights into the subcellular distribution of the protein .
Additional Applications
Some manufacturers have validated their SLC25A24 antibodies for additional applications:
-
ELISA: Proteintech's antibody (14669-1-AP) is validated for ELISA applications .
-
Indirect ELISA: The PBS formulation (14669-1-PBS) is specifically validated for this application .
These additional applications expand the utility of SLC25A24 antibodies for various research purposes, from protein quantification to high-throughput screening assays.
Cross-Reactivity and Specificity
The specificity and cross-reactivity profile of SLC25A24 antibodies are critical factors for experimental design and data interpretation, particularly in comparative studies involving multiple species.
Species Cross-Reactivity
Most commercial SLC25A24 antibodies demonstrate cross-reactivity with multiple mammalian species:
| Antibody | Tested Reactivity | Predicted Cross-Reactivity | Source |
|---|---|---|---|
| Proteintech 14669-1-AP/PBS | Human, mouse, rat | - | |
| Abcam ab221120 | Human | Mouse, rat, rabbit, cow (predicted) | |
| antibodies-online ABIN526192 | Human | - |
This cross-reactivity profile is advantageous for comparative studies across species and translational research that bridges animal models and human applications .
Specificity Validation
Manufacturers employ various methods to validate the specificity of their SLC25A24 antibodies:
-
Western blot analysis showing bands at the expected molecular weight (approximately 48 kDa) .
-
Positive controls in immunohistochemistry and immunofluorescence showing expected localization patterns .
-
Testing in multiple cell lines and tissue types to confirm consistent results .
The comprehensive validation data provided by manufacturers ensures that researchers can select antibodies with appropriate specificity for their experimental systems and research questions .
Handling Precautions
To maintain optimal antibody performance, manufacturers recommend:
-
Aliquoting the antibody upon receipt to minimize freeze-thaw cycles (especially for PBS-only formulations) .
-
Thawing frozen antibodies completely before use and mixing gently to ensure homogeneity .
-
Using appropriate personal protective equipment when handling antibodies containing preservatives like sodium azide .
-
Following manufacturer-specific protocols for each application to achieve optimal results .
Adhering to these storage and handling guidelines helps ensure consistent antibody performance across experiments and maximizes the useful life of these valuable reagents .
Research Applications and Significance
SLC25A24 antibodies have contributed significantly to our understanding of mitochondrial carrier proteins and their roles in cellular physiology and pathology.
Mitochondrial Transport Studies
SLC25A24 antibodies have been instrumental in elucidating the role of this protein as an adenyl nucleotide antiporter in the inner mitochondrial membrane . Research has revealed its function in transporting ATP-magnesium and phosphate across the mitochondrial membrane, contributing to the regulation of mitochondrial adenyl nucleotide pools .
Cellular Protection Mechanisms
Studies utilizing SLC25A24 antibodies have helped identify its protective role against oxidative stress-induced cell death . The protein appears to buffer calcium levels in the mitochondrial matrix through the formation of calcium-phosphate precipitates, which may be a critical mechanism for cellular survival under stress conditions .
Potential Clinical Relevance
While the search results don't explicitly mention clinical applications, the fundamental role of SLC25A24 in mitochondrial function and cellular protection suggests potential relevance to:
-
Mitochondrial disorders and diseases associated with energetic dysfunction .
-
Cellular responses to oxidative stress, which is implicated in numerous pathological conditions including neurodegenerative diseases and aging .
-
Metabolic disorders where mitochondrial adenyl nucleotide transport might be compromised .
The continued development and refinement of SLC25A24 antibodies will likely contribute to further discoveries regarding the role of this protein in both normal physiology and disease states.
Product Specs
Target Background
- SLC25A24 mutations induce a gain of pathological function. PMID: 29100093
- SLC25A24 mutations lead to impaired mitochondrial ATP synthesis. PMID: 29100094
- Genetic analyses in human and mouse models have revealed the importance of SLC25A24/Slc25a24 in regulating body fat mass and adipogenesis. PMID: 25599384
- N-terminal EF hands form a self-sequestered compact calmodulin-like Ca-sensor in the presence of Ca2+. PMID: 24332718
- SCaMC-1 is the human orthologue of the rabbit Efinal protein, which was reported to be located in peroxisomes. PMID: 15054102
- Three isoforms of the mitochondrial ATP-Mg/Pi carrier, APC1, APC2, and APC3, have been identified. They are most likely responsible for the net uptake or efflux of adenine nucleotides into or from the mitochondria. PMID: 15123600
Q&A
What is SLC25A24 and what cellular functions does it perform?
SLC25A24 is a mitochondrial inner membrane carrier protein that mediates the exchange of ATP-Mg2+ for phosphate (HPO42-) in a calcium-dependent manner. It is also known as short Ca2+-binding mitochondrial carrier 1 (SCaMC-1) or APC1 . The protein consists of an N-terminal calcium-binding domain containing four EF-hand motifs, followed by six transmembrane helices and a short C terminus . SLC25A24 plays a crucial role in mitochondrial function, particularly in resistance to oxidative stress . Functionally, it regulates ATP content in the mitochondrial matrix and influences mitochondrial membrane potential under oxidative stress conditions . Additionally, SLC25A24 has been implicated in fat-tissue metabolism, with knockout studies demonstrating an obesity-resistant phenotype in mice .
What are the validated applications for SLC25A24 antibodies?
SLC25A24 antibodies have been validated for multiple research applications:
It's important to note that these applications require specific optimization based on your experimental system. For IHC applications, antigen retrieval with TE buffer pH 9.0 is suggested, though citrate buffer pH 6.0 may be used as an alternative .
What molecular weight should be expected when detecting SLC25A24 in Western blots?
When performing Western blot analysis for SLC25A24, researchers should be aware of potential discrepancies between calculated and observed molecular weights:
This discrepancy between calculated and observed molecular weights is not uncommon in protein research. The mobility of proteins during gel electrophoresis can be affected by multiple factors including post-translational modifications, protein folding characteristics, and the experimental conditions . If you observe multiple bands, this may indicate the presence of different isoforms or differentially modified forms of the protein .
What species reactivity has been confirmed for commercial SLC25A24 antibodies?
Based on validation data, commercial SLC25A24 antibodies demonstrate cross-reactivity with samples from multiple species:
| Antibody | Confirmed Species Reactivity |
|---|---|
| Proteintech (14669-1-AP) | Human, mouse, rat |
| Elabscience (E-AB-63633) | Human, mouse, rat |
This cross-reactivity is particularly valuable for comparative studies across different model systems. The conservation of epitopes across species suggests functional importance of these regions within the SLC25A24 protein structure .
What are the recommended storage conditions for SLC25A24 antibodies?
Proper storage is critical for maintaining antibody activity and specificity:
Proper storage ensures antibody stability and prevents degradation that could compromise experimental results. When receiving a new shipment, it's advisable to immediately store the antibody at the recommended temperature .
How can SLC25A24 antibodies be optimized for detecting mitochondrial dysfunction?
When investigating mitochondrial dysfunction using SLC25A24 antibodies, several methodological considerations are crucial:
For Western blot analysis, subcellular fractionation to isolate mitochondrial proteins improves detection specificity. Use mitochondrial markers (e.g., VDAC or COX IV) as loading controls rather than traditional housekeeping proteins . When detecting changes in SLC25A24 expression under conditions of oxidative stress, time-course experiments are recommended as the protein levels may remain stable while its function changes .
For immunofluorescence studies, co-staining with mitochondrial markers (MitoTracker) allows assessment of SLC25A24 localization changes during mitochondrial swelling or ballooning . When studying the relationship between SLC25A24 and mitochondrial membrane potential, combine antibody detection with membrane potential-sensitive dyes and correlate antibody signal intensity with functional changes .
Research indicates that fibroblasts from individuals with mutations in SLC25A24 show mitochondrial swelling, which is exacerbated upon treatment with hydrogen peroxide (H₂O₂) . Therefore, when using SLC25A24 antibodies to assess mitochondrial dysfunction, parallel functional assays including ATP content measurement and membrane potential assessment provide a more comprehensive analysis .
What controls are essential when studying SLC25A24 mutations with antibody-based methods?
When investigating SLC25A24 mutations (such as the p.Arg217His mutation associated with craniosynostosis syndrome), the following controls are essential:
-
Antibody validation controls: Include wild-type cells/tissues and known positive samples (e.g., HEK-293 cells, mouse heart tissue) to confirm antibody specificity and optimize signal-to-noise ratio .
-
Expression level controls: Perform qPCR to assess mRNA levels parallel to protein detection, as mutations may affect transcript stability without altering protein levels. In previous studies with SLC25A24 mutations, cDNA sequencing and quantitative PCR showed stability of the mutant transcript .
-
Protein stability controls: Western blot analysis comparing wild-type and mutant SLC25A24 protein levels is critical, as research has shown that certain mutations (p.Arg217His) do not affect protein stability .
-
Functional controls: Include oxidative stress induction (e.g., H₂O₂ treatment) as research shows differential responses between wild-type and mutant SLC25A24 carriers under these conditions .
-
Co-localization controls: For microscopy studies, use mitochondrial markers to assess potential alterations in SLC25A24 localization due to mutations .
These controls help distinguish between mutation effects on protein expression versus functional alterations in the carrier activity, providing more comprehensive insights into the pathophysiological mechanisms.
How can discrepancies between predicted and observed molecular weights of SLC25A24 be resolved?
The discrepancy between calculated (51-53 kDa) and observed (48-50 kDa) molecular weights of SLC25A24 requires methodological approaches to ensure accurate protein identification:
-
Validation through multiple antibodies: Use antibodies targeting different epitopes of SLC25A24 to confirm the identity of the observed band .
-
Mass spectrometry confirmation: After immunoprecipitation or band excision from gels, perform mass spectrometry to definitively identify the protein and potential modifications .
-
Deglycosylation or dephosphorylation assays: Treat samples with appropriate enzymes to determine if post-translational modifications contribute to mobility shifts .
-
Genetic manipulation approaches: Include samples from SLC25A24 knockdown/knockout cells or overexpression systems as controls to confirm band identity .
-
Gradient gel systems: Utilize gradient gels for better resolution of proteins in the 40-60 kDa range to more accurately assess molecular weight .
When reporting results, acknowledge the discrepancy between calculated and observed molecular weights and provide evidence supporting the identification of SLC25A24 through the methodological approaches outlined above .
What considerations should be made when using SLC25A24 antibodies to study oxidative stress resistance?
When investigating SLC25A24's role in oxidative stress resistance, several methodological considerations enhance experimental rigor:
-
Gradual stress induction: Employ graduated concentrations of oxidative stressors (e.g., 1-10 μM H₂O₂) as research shows differential responses at varying concentrations .
-
Time-course analysis: Monitor SLC25A24 localization and expression at multiple time points after stress induction, as dynamic changes occur during the stress response .
-
Complementary functional assays: Combine antibody-based detection with measurements of:
-
Comparative analysis across cell types: Different cell types exhibit varying sensitivity to oxidative stress, influencing SLC25A24 function. Previous research comparing fibroblasts from affected individuals to control cells revealed important functional differences .
-
Knockdown/overexpression validation: Include SLC25A24 knockdown cells as previous work indicated that "knockdown of SLC25A24 in cancer cells was associated with increased cell death and mitochondrial swelling after treatment with hydrogen peroxide" .
This methodological approach allows researchers to distinguish between correlation and causation regarding SLC25A24's role in oxidative stress resistance.
How can SLC25A24 antibodies be integrated into studies of mitochondrial calcium homeostasis?
SLC25A24 functions as a calcium-dependent mitochondrial carrier, making it a valuable target for investigating mitochondrial calcium homeostasis:
-
Co-immunoprecipitation studies: Use SLC25A24 antibodies for co-IP to identify calcium-dependent interaction partners, providing insights into its regulatory networks .
-
Calcium chelation experiments: Compare SLC25A24 function/localization in normal and calcium-depleted conditions using chelators like BAPTA-AM, as the protein contains calcium-binding EF-hand motifs .
-
Site-directed mutagenesis approach: Use antibodies to detect expression/localization of SLC25A24 variants with mutations in the EF-hand domains to understand calcium binding effects .
-
Combined calcium imaging: Pair antibody-based detection with calcium-sensitive fluorescent probes to correlate SLC25A24 expression with mitochondrial calcium levels .
-
Super-resolution microscopy: Apply advanced imaging techniques with SLC25A24 antibodies to visualize its distribution within mitochondrial subcompartments relative to calcium microdomains .
These approaches help elucidate how SLC25A24 integrates calcium signaling with ATP-Mg²⁺/phosphate exchange across the mitochondrial inner membrane, a critical process for understanding mitochondrial energetics and stress responses.
What methodological approaches can assess SLC25A24's role in metabolic disorders using antibody-based techniques?
Given SLC25A24's implications in fat-tissue metabolism and obesity resistance, the following methodological approaches are valuable:
-
Tissue-specific expression analysis: Compare SLC25A24 protein levels across multiple metabolic tissues (adipose, liver, muscle) using optimized antibody dilutions for each tissue type .
-
Diet intervention studies: Analyze SLC25A24 expression changes in response to different dietary conditions, as research indicates "Slc25a24 expression was increased in white adipose tissue under a high-fat diet in WT mice" .
-
Metabolic challenge time-course: Monitor SLC25A24 expression/localization during glucose tolerance tests or insulin challenge using both antibody detection and functional metabolic parameters .
-
Co-localization with metabolic sensors: Combine SLC25A24 antibody detection with staining for metabolic sensors (e.g., AMPK, mTOR) to understand its integration with cellular energy sensing pathways .
-
Correlation with mitochondrial bioenergetics: Pair antibody-based detection with Seahorse analysis of mitochondrial respiration to connect SLC25A24 levels with functional bioenergetic parameters .
These approaches help mechanistically connect SLC25A24's mitochondrial carrier function to broader metabolic processes, potentially revealing therapeutic targets for metabolic disorders.
